1. Field
This disclosure is concerned generally with the use of alcohols as virucidal agents and is concerned specifically with the use of intermediate length alcohols to inactivate viruses in an aqueous solution of biologically active proteins.
2. Prior Art
The importance of eliminating viral infectivity in therapeutic products has long been recognized. This is especially true in the case of biologically active products derived from human blood or, more recently, from cell cultures used to make products of biotechnology (e.g. recombinant DNA products and monoclonal antibodies).
In considering virucidal agents for biologically active proteins, the primary goals are to assure complete virucidal action while not adversely affecting the biological activity of the protein. These goals require consideration of such variables as the protein itself, the nature of its activity and/or activity site, the virucidal agent, the importance and/or ease of its removal after use, and variables of the treatment itself, such as time, temperature, concentration, pH and ionic environment.
Although heat treatment alone can be used for virucidal treatment of some proteins (e.g. pasteurization conditions of at least 60.degree. C. for at least 10 hours for albumin or pasteurization of lyophilized forms of some proteins such as factor VIII), it is difficult in many cases to avoid loss of biological activity or utility when heat alone is used.
Recent examples of the use of heat against viruses in a lyophilized biologically active product (dry heat treatment) can be seen in U.S. Pat. No. 4,556,558, to A. Rubenstein. The use of heat against viruses in a stabilized aqueous solution (wet heat treatment) can be found in U.S. Pat. No. 4,440,679, to P. Fernandez and J. Lundblad.
To avoid some of the disadvantages or activity losses resulting from the use of heat alone, various chemical agents have been used or proposed as virucidal agents for biologically active proteins. See for example, U.S. Pat. No. 4,540,573, to A. Neurath and B. Horowitz, disclosing the use of compounds such as tri-n-butyl phosphate (TNBP) as virucidal agents for protein products.
The use of certain alcohols (chain lengths carbons) to inactivate lipid-containing viruses is disclosed by W. Snipes et al in Antimicrobial Agents and Chemotherapy, "Inactivation of Lipid-Containing Viruses by Long Chain Alcohols", Vol. 11, No. 1, pp. 98-104, Jan. 1977. However, those studies were not concerned with preparations of biologically active proteins or the need to maintain the activity of such proteins. Although other alcohols were used, the authors suggest the use of alcohols that are extremely insoluble in aqueous media such as C10 to C14 alcohols. Because of the low water solubility, the preferred alcohols first had to be prepared in 95% ethanol at 100 times the desired final concentration. The final treatment was in solutions adjusted to a pH of at least 7.2. Although the authors used alcohols ranging from C4 to C.sub.18, they point out that a striking peak in virucidal activity was found for saturated alcohols having chain lengths from 10 to 14 carbons.
Although the C.sub.10 - 14 alcohols were shown to have good virucidal action, their relatively low water solubility is a disadvantage in view of the initial preparation steps required. We have found this disadvantage can be avoided and that a class of alcohols can now be used to treat biologically active proteins under controlled pH conditions which in most cases has little adverse effect of biological activity. Details of our method and preferred embodiments are described below.